PROJECT SUMMARY/ABSTRACT Acinetobacter baumannii (Ab) is an opportunistic bacterium has an alarming predisposition to attain multi-drug resistance (MDR), and infections with MDR-Ab strains are linked to greater morbidity and mortality. Ab is categorized by the World Health Organization as a critical priority for the research and development of new antimicrobial therapies. Plasmids serve as vehicles for the spread of MDR among Ab clinical isolates, and type IV secretion system (T4SS)-dependent conjugation is an effective mean by which many of these plasmids are disseminated. Acinetobacter contains three classes of plasmids: the Large Conjugative Plasmid (LCP), the Medium-sized Conjugative Plasmid (MCP), and the small Mobilizable Plasmid (SMP) families. These plasmids contain conserved T4SS machinery and/or elements that target them for mobilization, supporting that conjugation plays a critical role in their dissemination. Successful T4SS-mediated conjugation requires close contact between the plasmid ?donor? strain and the plasmid ?recipient? strain, as well as the subsequent survival of the recipient. This is contrasted with the function of the Type-VI secretion system (T6SS), which mediates indiscriminate, contact-dependent killing of neighboring, non-sister bacterial competitors. Most Ab strains carry a constitutively active T6SS. This poses a unique challenge to conjugative plasmids, as Ab plasmid ?donors? can kill potential ?recipients? and vice versa. However, Acinetobacter plasmids are successfully disseminating, suggesting that they have evolved mechanisms to overcome this hypothetical restriction. This proposal addresses the relationship between T6SS and plasmid conjugation in Ab. We previously demonstrated that some LCPs have the unique ability to abrogate the expression of the T6SS encoded in the host chromosome. Thus, we hypothesize that silencing the T6SS is essential for plasmid conjugation and the resulting dissemination of multidrug resistance among Acinetobacter. Furthermore, we propose that in the absence of their own conjugative machinery, horizontal transfer of SMPs relies on LCP- encoded conjugation machineries and LCP-mediated T6SS repression. Because potential plasmid recipients may employ their T6SS to kill potential plasmid donors, we propose that a constitutively active T6SS in Acinetobacter can confer immunity against conjugation. Finally, we hypothesize that the role of conjugative plasmids in MDR dissemination among Ab clinical isolates has gone largely unnoticed. This is likely due to the limited understanding of Acinetobacter plasmid biology and the lack of a systematic epidemiological study of their prevalence. Therefore, we propose a pilot prospective analysis to determine the incidence of conjugative plasmids among contemporary Acinetobacter clinical isolates, their contribution to MDR, and their effects on T6SS activity. This proposal may open new avenues to investigate innovative drugs with capacity of eliminating Ab MDR-plasmids and re-sensitizing pathogenic strains to available antibiotics. These ?antiplasmid? molecules would have a tremendous clinical impact in the treatment of Acinetobacter MDR infections.